Influenza A virus (IAV) and especially the highly pathogenic IAV strains, such as those causing the recent H5N1 and H1N1 outbreaks, represent a significant public health concern. These strains of IAV cause significantly greater morbidity and mortality than seasonal IAV strains. However, the exact cause of the increased morbidity and mortality remains controversial, with increased viral pathogenicity, increased immunopathology, and increase co-infections proposed as possible mechanisms. This proposal will examine the early host response elements to IAV infection, specifically examining the role that mast cells and protease- activated receptors play in regulating the morbidity and mortality associated with IAV infection. Mast cells could be critical in orchestrating the host response to respiratory infection with IAV, because they are found at elevated frequencies around the bronchi within the lungs. Furthermore, our preliminary data show that mast cell-deficient mice develop significantly less IAV-association morbidity. Therefore, we propose to determine how mast cells are activated during respiratory infection with IAV and what role they play in regulating disease outcome. Secondly, we will determine how mast cells are orchestrating the deleterious host inflammatory response following IAV infection. Specifically, mast cell activation of the protease-activated receptor family on alveolar epithelial cells will be explored, as well as the subsequent regulation of inflammatory chemokines and cytokines by both mast cells and the protease-activated receptor family. Our preliminary data support the hypothesis that mast cells and the protease-activated receptor family are important in regulating morbidity associated with respiratory IAV infection. Thus, by understanding these early events, we hope to identify novel pharmacological targets, which can be used to limit morbidity and mortality from IAV infection. As can be seen, the focus of this proposal is quite distinct from the previous research that I have conducted. However, I have been well trained in the experimental systems and techniques needed for the successful completion of the proposed studies. For example, I have gained extensive experimental experience in the use of mice for understanding the immune response to numerous viral pathogens, including IAV. In addition, I have gained substantial training in many of the techniques required for the successful completion of the proposed studies, which include multi-parameter flow cytometry (we are routinely conducting 12 parameter studies now), primary cell culture from bone marrow, Luminex" bead assays for the detection of multiple cytokines/chemokines at the same time, viral plaque assays, adoptive cellular transfers, and ELISA assays. Together, this previous experience provides a strong experimental foundation, which will allow me the flexibility to apply this knowledge and technical ability to a distinct area of study, therefore developing a new and independent research direction that addresses critical gaps in knowledge of immune regulation of viral infection as an Assistant Professor. Furthermore, my current and future research institutes provide me ample technical opportunities to conduct these studies, including access to a 10-color BD LSRII cytometer, a g-cell irradiator for making bone marrow chimeras, and BioPlex200 machine for running Luminex plates. Additionally, faculty members at both my current and future universities provide a sound network of knowledge which will be invaluable for the successfully completion of the proposed studies. Each year, influenza A virus infection leads to significant morbidity and mortality within certain groups, while periodic pandemic strains cause significantly greater morbidity and mortality throughout the entire population. What causes the significantly elevated rates of morbidity and mortality remains controversial. This proposal will examine the early events during the host response to influenza A virus infection and could elucidate potential novel therapeutic targets for limiting morbidity and mortality associated with influenza A virus infection.